Dysregulation of Wnt signaling in bone of type 2 diabetes mellitus and diabetic Charcot arthropathy.

BACKGROUND: Type 2 diabetes mellitus (T2DM) patients show a markedly higher fracture risk and impaired fracture healing when compared to non-diabetic patients. However in contrast to type 1 diabetes mellitus, bone mineral density in T2DM is known to be normal or even regionally elevated, also known as diabetic bone disease. Charcot arthropathy is a severe and challenging complication leading to bone destruction and mutilating bone deformities. Wnt signaling is involved in increasing bone mineral density, bone homeostasis and apoptotic processes. It has been shown that type 2 diabetes mellitus is strongly associated with gene variants of the Wnt signaling pathway, specifically polymorphisms of TCF7L2 (transcription factor 7 like 2), which is an effector transcription factor of this pathway. METHODS: Bone samples of 19 T2DM patients and 7 T2DM patients with additional Charcot arthropathy were compared to 19 non-diabetic controls. qPCR analysis for selected members of the Wnt-signaling pathway (WNT3A, WNT5A, catenin beta, TCF7L2) and bone gamma-carboxyglutamate (BGLAP, Osteocalcin) was performed and analyzed using the 2-ΔΔCt- Method. Statistical analysis comprised one-way analysis of variance (ANOVA). RESULTS: In T2DM patients who had developed Charcot arthropathy WNT3A and WNT5A gene expression was down-regulated by 89 and 58% compared to healthy controls (p < 0.0001). TCF7L2 gene expression showed a significant reduction by 63% (p < 0.0001) and 18% (p = 0.0136) in diabetic Charcot arthropathy. In all diabetic patients BGLAP (Osteocalcin) was significantly decreased by at least 59% (p = 0.0019). CONCLUSIONS: For the first time with this study downregulation of members of the Wnt-signaling pathway has been shown in the bone of diabetic patients with and without Charcot arthropathy. This may serve as future therapeutic target for this severe disease.

Pantoprazole decreases cell viability and function of human osteoclasts in vitro.

Proton pump inhibitors (PPIs) are commonly prescribed drugs that decrease stomach acidity and are thus often used to treat gastroesophageal reflux disease and as a preventative agent for the adverse effects of nonsteroidal anti-inflammatory drugs on the stomach mucosa. In recently published literature, an association between proton pump inhibitor administration and increased fracture risk has been stated. In order to reveal the underlying pathomechanisms of these observations, the effects of pantoprazole, a representative of the proton pump inhibitors, on human osteoclasts in vitro were evaluated in this study. Osteoclasts were stimulated with increasing concentrations of pantoprazole ranging from 0 µg/mL to 10 µg/mL over a period of seven days. Cell viability and tartrate-resistant acid phosphatase (TRAP) activity assays were performed after 1 day, 3 days, and 7 days, respectively. Here, stimulated osteoclasts presented a significantly lower viability and TRAP activity than the negative controls. Osteoclast-specific gene expression was evaluated after seven days and revealed no significant differences between all samples. Overall, the bone degrading and resorptive function of osteoclasts is inhibited by the administration of proton pump inhibitors. While PPI-related fractures through "basic multicellular unit" dysfunction are unlikely, the underlying pathomechanism remains unknown.

Pantoprazole increases cell viability and function of primary human osteoblasts in vitro.

Proton pump inhibitors (PPIs) are a class of drugs that irreversibly inhibit the H(+)/K(+)-ATPase in gastric parietal cells. Since an association between PPI use and increased fracture risk has been found, the aim of this study was to detect potential adverse effects of pantoprazole, a representative of the PPIs, on primary human osteoblasts in vitro. The isolated cells were stimulated with pantoprazole concentrations ranging from 0 µg/ml to 10 µg/ml. Changes in proliferation, total cell number, viability, cytotoxicity, alkaline phosphatase activity, total protein synthesis and gene expression on mRNA level were determined over a period of 7 days. Pantoprazole stimulation resulted in increased viability and decreased cytotoxicity in the osteoblasts. The proliferation rate was stable and so was the relative cell number. Only at the highest pantoprazole concentration on day 7, a slight decrease of the cell number was detected. Alkaline phosphatase activity increased over the tested period under exposure to pantoprazole (p < 0.05 at 3 µg/ml and 10 µg/ml pantoprazole). Osteoblast-specific gene expression was increased through pantoprazole stimulation compared to the control on day 3. Towards day 7, gene expression returned to baseline levels or decreased slightly compared to unexposed cells. Interestingly, this in vitro experiment detected no evidence of adverse effects of PPIs on primary human osteoblasts. Osteoblasts were rather more viable with increased mitochondrial activity, gene expression and protein synthesis under pantoprazole stimulation. Therefore, these in vitro results do not suggest that impaired osteoblast function is the cause of an increased fracture risk in patients under PPI therapy.